Difference between revisions of "Part:BBa K5322030"
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| − | In the context of intestinal inflammation, the presence of CD8+ T cells often leads to an immune hyperreactivity. To downregulate the immune response at the site of inflammation, we plan to utilize the PD-1/PD-L1 immune checkpoint to inhibit the activity of CD8+ cytotoxic T cells. For the purpose of facilitating further experiments, we selected mouse PD-L1 (BBa_K5322033) and conducted codon optimization, designing the plasmid pET29a-J23119-RBS-PD-L1 (Mus)-T7, as illustrated in the figure below. | + | In the context of intestinal inflammation, the presence of CD8<sup>+</sup> T cells often leads to an immune hyperreactivity. To downregulate the immune response at the site of inflammation, we plan to utilize the PD-1/PD-L1 immune checkpoint to inhibit the activity of CD8+ cytotoxic T cells. For the purpose of facilitating further experiments, we selected mouse PD-L1 (BBa_K5322033) and conducted codon optimization, designing the plasmid pET29a-J23119-RBS-PD-L1 (Mus)-T7, as illustrated in the figure below. |
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==Construction of the plasmid== | ==Construction of the plasmid== | ||
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| − | The sequence of J23119-RBS-PD-L1 (Mus)-T7 was sent to a company for synthesis. After two weeks, the company returned results indicating that random mutations occurred during the process of recombining the fragment with the vector and introducing it into Escherichia coli. Given the uncertainties associated with these random mutations, we decided to abandon this approach. | + | The sequence of J23119-RBS-PD-L1 (Mus)-T7 was sent to a company for synthesis. After two weeks, the company returned results indicating that random mutations occurred during the process of recombining the fragment with the vector and introducing it into <i>Escherichia coli</i>. Given the uncertainties associated with these random mutations, we decided to abandon this approach. |
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Latest revision as of 12:11, 2 October 2024
Programmed Cell Death 1 Ligand 1 [Mus musculus] Expression System
Contents
Usage and Biology
In the context of intestinal inflammation, the presence of CD8+ T cells often leads to an immune hyperreactivity. To downregulate the immune response at the site of inflammation, we plan to utilize the PD-1/PD-L1 immune checkpoint to inhibit the activity of CD8+ cytotoxic T cells. For the purpose of facilitating further experiments, we selected mouse PD-L1 (BBa_K5322033) and conducted codon optimization, designing the plasmid pET29a-J23119-RBS-PD-L1 (Mus)-T7, as illustrated in the figure below.
Figure 1-1 Plasmid pET29a-J23119-RBS-[PD-L1(Mus)]-T7
Construction of the plasmid
The sequence of J23119-RBS-PD-L1 (Mus)-T7 was sent to a company for synthesis. After two weeks, the company returned results indicating that random mutations occurred during the process of recombining the fragment with the vector and introducing it into Escherichia coli. Given the uncertainties associated with these random mutations, we decided to abandon this approach.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 356
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal NheI site found at 314
Illegal NheI site found at 939
Illegal PstI site found at 356 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 205
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 356
- 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 356
Illegal NgoMIV site found at 745
Illegal AgeI site found at 577 - 1000COMPATIBLE WITH RFC[1000]